LOCUS       62056_13065        4705 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4705)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..4705
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        97..131
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        149..224
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(257..273)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(332..348)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(356..372)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(380..410)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(425..446)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(734..1322)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1496..2284)
                     /codon_start=1
                     /label=SmR
                     /note="aminoglycoside adenylyltransferase (Murphy, 1985)"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(2285..2389)
                     /label=AmpR promoter
     primer_bind     2863..2879
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2917..2933
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2998..4416)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
ORIGIN
        1 cgggcctaaa gaactaaaaa atctatctgt ttcttttcat tctctgtatt ttttatagtt
       61 tctgttgcat gggcataaag ttgcaagctt gatatcttga cagctagctc agtcctaggt
      121 ataatactag tcgagaccat tggtctcagt tttagagcta gaaatagcaa gttaaaataa
      181 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt tgatatcgaa
      241 ttcctgcagc ccgggggatc cactagttct agagcggccg ccaccgcggt ggagctcatc
      301 ccaatggcgc gccgagcttg gctcgagcat ggtcatagct gtttcctgtg tgaaattgtt
      361 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
      421 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
      481 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
      541 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
      601 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      661 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      721 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      781 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      841 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      901 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      961 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     1021 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     1081 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     1141 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     1201 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     1261 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     1321 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     1381 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     1441 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttatttgc
     1501 cgactacctt ggtgatctcg cctttcacgt agtggacaaa ttcttccaac tgatctgcgc
     1561 gcgaggccaa gcgatcttct tcttgtccaa gataagcctg tctagcttca agtatgacgg
     1621 gctgatactg ggccggcagg cgctccattg cccagtcggc agcgacatcc ttcggcgcga
     1681 ttttgccggt tactgcgctg taccaaatgc gggacaacgt aagcactaca tttcgctcat
     1741 cgccagccca gtcgggcggc gagttccata gcgttaaggt ttcatttagc gcctcaaata
     1801 gatcctgttc aggaaccgga tcaaagagtt cctccgccgc tggacctacc aaggcaacgc
     1861 tatgttctct tgcttttgtc agcaagatag ccagatcaat gtcgatcgtg gctggctcga
     1921 agatacctgc aagaatgtca ttgcgctgcc attctccaaa ttgcagttcg cgcttagctg
     1981 gataacgcca cggaatgatg tcgtcgtgca caacaatggt gacttctaca gcgcggagaa
     2041 tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc gttgatcaaa gctcgccgcg
     2101 ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc aatatcactg tgtggcttca
     2161 ggccgccatc cactgcggag ccgtacaaat gtacggccag caacgtcggt tcgagatggc
     2221 gctcgatgac gccaactacc tctgatagtt gagtcgatac ttcggcgatc accgcttccc
     2281 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     2341 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     2401 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     2461 ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac
     2521 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     2581 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat
     2641 cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa
     2701 ggagaaaata ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     2761 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
     2821 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     2881 aattgacgcg tattgggatg gtaccgggcc ccccctcgag gtcgacggta tcgatacggc
     2941 atcagagcag attgtactga gagtgcacca taatcggcat tttcttttgc gtttttattt
     3001 gttaactgtt aattgtcctt gttcaaggat gctgtctttg acaacagatg ttttcttgcc
     3061 tttgatgttc agcaggaagc taggcgcaaa cgttgattgt ttgtctgcgt agaatcctct
     3121 gtttgtcata tagcttgtaa tcacgacatt gtttcctttc gcttgaggta cagcgaagtg
     3181 tgagtaagta aaggttacat cgttaggatc aagatccatt tttaacacaa ggccagtttt
     3241 gttcagcggc ttgtatgggc cagttaaaga attagaaaca taaccaagca tgtaaatatc
     3301 gttagacgta atgccgtcaa tcgtcatttt tgatccgcgg gagtcagtga acagatacca
     3361 tttgccgttc attttaaaga cgttcgcgcg ttcaatttca tctgttactg tgttagatgc
     3421 aatcagcggt ttcatcactt ttttcagtgt gtaatcatcg tttagctcaa tcataccgag
     3481 agcgccgttt gctaactcag ccgtgcgttt tttatcgctt tgcagaagtt tttgactttc
     3541 ttgacggaag aatgatgtgc ttttgccata gtatgctttg ttaaataaag attcttcgcc
     3601 ttggtagcca tcttcagttc cagtgtttgc ttcaaatact aagtatttgt ggcctttatc
     3661 ttctacgtag tgaggatctc tcagcgtatg gttgtcgcct gagctgtagt tgccttcatc
     3721 gatgaactgc tgtacatttt gatacgtttt tccgtcaccg tcaaagattg atttataatc
     3781 ctctacaccg ttgatgttca aagagctgtc tgatgctgat acgttaactt gtgcagttgt
     3841 cagtgtttgt ttgccgtaat gtttaccgga gaaatcagtg tagaataaac ggatttttcc
     3901 gtcagatgta aatgtggctg aacctgacca ttcttgtgtt tggtctttta ggatagaatc
     3961 atttgcatcg aatttgtcgc tgtctttaaa gacgcggcca gcgtttttcc agctgtcaat
     4021 agaagtttcg ccgacttttt gatagaacat gtaaatcgat gtgtcatccg catttttagg
     4081 atctccggct aatgcaaaga cgatgtggta gccgtgatag tttgcgacag tgccgtcagc
     4141 gttttgtaat ggccagctgt cccaaacgtc caggcctttt gcagaagaga tatttttaat
     4201 tgtggacgaa tcgaactcag gaacttgata tttttcattt ttttgctgtt cagggatttg
     4261 cagcatatca tggcgtgtaa tatgggaaat gccgtatgtt tccttatatg gcttttggtt
     4321 cgtttctttc gcaaacgctt gagttgcgcc tcctgccagc agtgcggtag taaaggttaa
     4381 tactgttgct tgttttgcaa actttttgat gttcatcgtt catgtctcct tttttatgta
     4441 ctgtgttagc ggtctgcttc ttccagccct cctgtttgaa gatggcaagt tagttacgca
     4501 caataaaaaa agacctaaaa tatgtaaggg gtgacgccaa agtatacact ttgcccttta
     4561 cacattttag gtcttgcctg ctttatcagt aacaaacccg cgcgatttac ttttcgacct
     4621 cattctatta gactctcgtt tggattgcaa ctggtctatt ttcctctttt gtttgataga
     4681 aaatcataaa aggatttgca gacta
//
